Treatment of autoimmune disorders with oral interferon

ABSTRACT

Autoimmune disorders are treated by the administration of human interferon, particularly IFN- alpha  or IFN- beta , at a dosage of from about 0.01 to about 5 IU/lb./day such that the interferon is held in contact with the patient&#39;s oral and pharyngeal mucosae. The interferon is administered in a solid dosage form, e.g., a saliva-dissolvable lozenge.

BACKGROUND OF THE INVENTION

This application is a divisional of U.S. patent application No.08/305,418, filed Sep. 13, 1994, which is a continuation of U.S. patentapplication No. 08/099,353, filed Jan. 26, 1993, now abandoned, which isa continuation of U.S. patent application No. 07/875,071, filed Apr. 28,1992, now abandoned, which is a continuation of U.S. patent applicationNo. 07/110.501, filed Oct. 26, 1987, now abandoned, which is acontinuation-in-part of U.S. patent application No. 06/927,834, filedNov. 6, 1986, now abandoned.

This invention relates generally to an improved method of treatingdiseases of immuno-pathologic etiology in warm-blooded vertebrates usinginterferon in low oral dosages. This invention also relates to the useof interferon in low oral dosages to potentiate disease-correctiveimmune responses in warm-blooded vertebrates afflicted withimmuno-resistant diseases characterized by apparent hyperactive orhypoactive immune system function.

"Interferon" is a term generically comprehending a group of vertebrateglycoproteins and proteins which are known to have various biologicalactivities, such as antiviral, antiproliferative, and immunomodulatoryactivity at least in the species of animal from which such substancesare derived. The following definition for interferon has been acceptedby an international committee assembled to devise a system for theorderly nomenclature of interferons: "To qualify as an interferon afactor must be a protein which exerts virus nonspecific, antiviralactivity at least in homologous cells through cellular metabolicprocesses involving synthesis of both RNA and protein." Journal ofInterferon Research, 1, pp. vi (1980). "Interferon" as used herein indescribing the present invention shall be deemed to have thatdefinition.

Since the first descriptions of interferon by Isaacs and Lindeman See,Proc. Roy. Soc. London (Ser. B), Vol. 147, pp. 258 et seg. (1957) andU.S. Pat. No. 3,699,222!, interferon has been the subject of intensiveresearch on a worldwide basis. The literature is replete withpublications concerning the synthesis of interferon, its proposedmolecular characterizations, its clinical applications and proposedmechanisms of its antitumor, antiviral, and immune system activities.

Because of the intensity and disparate origins of research concerninginterferon and its characteristics and uses, there exists a substantiallack of uniformity in such matters as classification of interferontypes. There are also numerous, sometimes contradictory, theoriesconcerning the mode of action of interferon in producing clinicaleffects.

Although originally isolated from cells of avian origin (chick allantoiccells), interferon production has been observed in cells of all classesof vertebrates, including mammals, amphibians, birds and reptiles.Interferon production by vertebrate cells is seldom spontaneous but isoften readily "induced" by treatment of cells (in vivo or in vitro) witha variety of substances including viruses, nucleic acids (includingthose of viral origin as well as synthetic polynucleotides),lipopolysaccharides, and various antigens and mitogens.

Interferons have generally been named in terms of the species of animalcells producing the substance (e.g., human, murine, or bovine), the typeof cell involved (e.g., leukocyte, lymphoblastoid, fibroblast) and,occasionally, the type of inducing material responsible for interferonproduction (e.g., virus, immune). Interferon has been loosely classifiedby some researchers according to induction mode as either Type I or TypeII, with the former classification comprehending viral and nucleic acidinduced interferon and the latter class including the material producedas a lymphokine through induction by antigens and mitogens. Morerecently, the international committee devising an orderly nomenclaturesystem for interferon has classified interferon into types on the basisof antigenic specificities. In this newer classification, thedesignations alpha (α), beta (β), and gamma (γ) have been used tocorrespond to previous designations of leukocyte, fibroblast, and typeII (immune) interferons, respectively. Alpha and beta interferons areusually acid-stable and correspond to what have been called type Iinterferons; gamma interferons are usually acid-stable and correspond towhat has been called type II interferons. The international committee'snomenclature recommendations apply only to human and murine interferons.Journal of Interferon Research, 1 pp. vi (1980).

In its earliest applications, interferon was employed exclusively as anantiviral agent and the most successful clinical therapeuticapplications to date have been in the treatment of viral orvirus-related disease states. It became apparent, however, thatexogenous interferon was sometimes capable of effecting regression orremission of various metastatic diseases. An overview of currentclinical trials of interferon as an antiviral and antiproliferativetherapeutic agent is contained in Interferon: In Vivo and ClinicalStudies, Volume 4, Eds: N. B. Finter and R. K. Oldham, Academic Press,New York, 1985.

The clinical agent of choice for the present invention is humanleukocyte interferon, "mass-produced" by procedures involving collectionand purification of vast quantities of human buffy coat leukocytes,induction with virus, and isolation from culture media.

In the work described above, interferon has been administeredparenterally, i.e., intramuscularly and intradermally, with somesuccessful topical and intranasal usages having been reported. It hasseldom been administered intravenously because of substantial adverseeffects attributable to "contaminants" in crude and even highly purifiedisolates.

As discussed above, there has been a significant research effortdirected to the evaluation of therapeutic effects of interferon for awide variety of diseases having an auto-immuno-pathologic basis. Beforeapplicant's first report of successful oral administration of interferonin his U.S. patent application Ser. No. 415,525 (now U.S. Pat. No.4,462,985), there was no recognition in the art of the potential offeredby oral administration of interferon. The generally held belief was thatinterferon could not survive the digestive conditions of the upperalimentary canal.

Since applicant's first disclosure of the immunotherapeutic benefitachievable via oral administration of interferon of heterologousmammalian species, he has continued to investigate the efficacy oforally administered interferon. In U.S. Pat. No. 4,497,795, issued Feb.5, 1985, applicant described and claimed the use of interferonadministered orally or via intravenous administration to stimulateappetite and feed efficiency of bovine and porcine species. Morerecently applicant has described in now pending U.S. applications theuse of interferon at dosages less than about 5 IU per pound of bodyweight for increasing feed efficiency and food utilization inwarm-blooded vertebrates, for preventing and treating shipping fever,and for enhancing vaccine efficiency.

Human alpha-interferon has been marketed under the trademark Agriferon®by Immunomodulator Laboratories, Inc. ("IML") of Stafford, Tex. forveterinary use in Texas since February 1985. The product is sold fororal administration to cattle to promote growth and feed efficiency andto prevent or treat viral respiratory infections. IML began selling analpha-interferon product for horses in 1986. Both products are soldunder a license of my U.S. Pat. No. 4,462,985.

SUMMARY OF THE INVENTION

Interferon contacting the oral and/or pharyngeal mucosa, in amounts ofless than 5 IU/lb of body weight per day is consistently effective topotentiate disease-corrective immune responses in vertebrates afflictedwith immuno-resistant disease states characterized by apparenthyperactive or hypoactive immune system function. Treatment inaccordance with the present invention has been shown to effect remissionof neoplastic disease, hyperallergenicity, immuno-resistant orimmuno-debilitating viral infections and autoimmune disorderscharacterized by chronic tissue degenerative inflammation.

DETAILED DESCRIPTION OF THE INVENTION

The clinical agent of choice for use in the present invention is humanleukocyte interferon (human alpha-interferon), "mass-produced" byprocedures involving collection and purification of quantities of humanbuffy coat leukocytes, induction of interferon production with virus,and isolation of culture media. (See "Preparation of HumanAlpha-Interferon" below.) Also acceptable for use in accordance withpresent intention are human alpha-interferon products produced byrecombinant DNA technology and now commercially available fromSchering-Plough (as Intron®) and Hoffmann-LaRoche (as Roferon®) andapproved by the FDA for treatment (parenterally) of hairy cell leukemiaof man. Such recombinant interferon products are believed to beparticularly effective when used in combination. Gamma interferon isalso available by recombinant technology and is presently undergoingclinical trials by Genentech and others. Fibroblast interferon(beta-interferon) can be prepared in accordance with Example 1 inapplicant's U.S. Pat. No. 4,462,985 issued Jul. 31, 1984, the disclosureof which is hereby expressly incorporated by reference.

Interferon of human and murine origins has been quantified in the art interms of International Units ("IU"). As used herein, a "unit" ofinterferon (to be distinguished from "IU") shall mean the reciprocal ofa dilution of interferon-containing material that, as determined byassay, inhibits one-half the number of plaques of a challenge virus, thechallenge virus being the vesicular stomatitis virus ("VSV"). Soquantified a "unit" of interferon is routinely found to be aboutone-tenth the quantity of interferon represented by one "IU." In otherwords, for the purpose of defining the present invention, 1 unit ≈0.1IU.

The present invention relates to an improved method of treatment ofimmuno-resistant disease states with interferon. The present inventionis directed to the treatment of diseases in warm-blooded vertebrates,particularly certain diseases which the immune system of many species ispoorly equipped to handle, as evidenced by either a lack of diseasedefeating response and/or an apparently misdirected immune responseresulting in a chronic tissue degenerative inflammatory condition orother physical complications. While there has been a significantresearch effort directed to the use of interferon for treatment of suchdiseases, reported results, although positive overall, have beeninconsistent. The principle reason for such inconsistency in view of mymost recent research efforts is that earlier investigators have failedto define optimum dosage and route of interferon administration.

The present invention is based on applicant's discovery that interferoncan be used as a consistently effective therapeutic agent for treatmentof diseases having an immunopathologic basis--characterized byinadequate immune response and persistence of the disease or by anapparent hyperactive immune response resulting in tissue degenerativeinflammatory conditions and related physical manifestations. Applicanthas found that interferon, contacting the oral and pharyngeal mucosa inamounts from about 0.01 to about 5 IU/lb of body weight per day, isconsistently efficacious for the treatment of diseases to which theimmune system of many warm-blooded vertebrates does not effectivelyrespond.

Disease conditions treated in accordance with the present inventioninclude apparent autoimmune disorders characterized by a chronic tissuedegenerative inflammatory condition. Diseases so characterized includemultiple sclerosis, rheumatoid arthritis, stomatitis, and lupuserythematosus. Treatment of such disease is in accordance with thepresent invention comprises administering interferon at a dosage of 0.01to about 5 IU/lb per day in a dosage form adapted to promote contact ofsaid dosage of interferon with the oral and pharyngeal mucosa of saidanimal. Preferably, the dosage of interferon is from 0.1 to about 4.0IU/lb per day, more preferably 0.5 to about 1.5 IU/lb of body weight perday. Alpha interferon, derived from tissue culture or by recombinant DNAtechniques, is a preferred therapeutic agent in accordance with thisinvention. Some data have indicated better efficacy, i.e., a morepronounced immunomodulatory effect, where the interferon is nothomologous to the species being treated. Alpha interferon can beadministered alone or in combination with beta interferon or gammainterferon.

It is critical that the interferon be administered in a dosage formadapted to assure maximum contact of the interferon in said dosage formwith the oral and pharyngeal mucosa of the human or animal undergoingtreatment. Contact of interferon with the mucosa can be enhanced bymaximizing residence time of the treatment solution in the oral orpharyngeal cavity. Thus, best results seem to be achieved in humanpatients when the patient is requested to hold said solution ofinterferon in the mouth for a period of time. Contact of interferon withthe oral and pharyngeal mucosa and thereafter with the lymphatic systemof the treated human or animal is unquestionably the most efficientmethod administering immunotherapeutic amounts of interferon.

Another disease condition responding to treatment in accordance with thepresent invention is neoplastic disease. Thus, the administration ofinterferon in accordance with the above description can, alone or incombination with other drugs or therapy, help effect remission ofcancers such as malignant lymphoma, melanoma, mesothelioma, Burkittlymphoma and nasopharyngeal carcinoma and other neoplastic diseases,especially those of known or suspected viral etiology. Based on theresults observed to date, it is believed that applicant's presentlydescribed method of treatment will similarly help effect remission ofHodgkin's Disease and leukemia.

Other disease conditions responding to treatment in accordance with thepresent invention are infectious diseases of viral origin in, forexample, human, avian, porcine, canine and feline species.Significantly, viral infection typically exhibiting persistentresistance to treatment have shown a dramatic response to treatment withinterferon in low doses contacting the oral and pharyngeal mucosa ofinfected patients. Beneficial results have been attained utilizing thepresent method to treat dogs having canine parvovirus and canineherpesvirus infections. Further, feline leukemia and feline infectiousperitonitis have been shown to be particularly susceptible to treatmentwith alpha interferon and beta interferon in accordance with thisinvention.

Exemplary of human viral infections showing remarkable response totreatment in accordance with the present invention are infections ofhuman rhinovirus (common cold), herpes simplex I virus (cold sores) andhuman papovavirus (warts). Based on treatment results to date, it isexpected that contact of interferon at low dosage with the oral andpharyngeal mucosa will provide an effective treatment for AcquiredImmune Deficiency Syndrome (AIDS) and disease conditions having theherpes simplex II virus as the causative agent. A patient experiencing acondition of viral myocarditis has responded favorably to the presenttreatment. Warts often dissipate within six to eight weeks afterinitiating treatment in accordance with this invention. Interferonadministration in accordance with this invention can also be used tohelp prevent viral infections, for example, infections by the causitiveagents of flus and colds, and to minimize the symptoms associated withsuch viral infections.

Other afflictions responding to contact of low dosage interferon arehyperallergenic conditions such as asthma. One "side effect" noted bypatients treated in accordance with this invention is improved skincomplexion. Thus, administration of interferon in dosages of about 0.01to about 5 IU/lb of body weight per day is effective to treat acne,specifically and improve human skin complexion generally.

Further, stimulating the immune system by oral contact with low dosageinterferon is believed to assist the body in fighting bacterialinfection. Treatment in accordance with this invention alone or incombination with therapeutic amounts of antibiotics can be especiallyeffective in knocking down infections of antibiotic resistantmicroorganisms.

Administration of interferon in accordance with the present invention ispreferably continued until the symptoms of the disease condition beingtreated subside. This can range from a period of one day, for example,where a human rhinovirus is the disease causative agent, to a period ofup to six months for treatment of neoplastic disease. Rheumatoidarthritis patients are pain free within 2 to 10 days of initiatingtreatment in accordance with the present invention. However, treatmentof that disease is preferably conducted by administration of interferonfor up to about three (3) months.

Daily dosage of interferon can be administered as a single dosage or,preferably, it is divided and administered in a multiple-dose dailyregimen. A staggered regimen of at least one, for example, one to threedays treatment per week or month, can be used as an alternative tocontinuous daily treatment.

Interferon can be administered in accordance with this invention ineither a liquid (solution) or solid dosage form. Thus interferon can beadministered dissolved in a buffered aqueous solution typicallycontaining a stabilizing amount (1-5% by weight) of blood serums.Exemplary of a buffered solution suitable as a carrier of interferonadministered in accordance with this invention is phosphate bufferedsaline prepared as follows:

A concentrated (20×) solution of phosphate buffered saline (PBS) wasprepared by dissolving the following reagents in sufficient water tomake 1,000 ml of solution: sodium chloride, 160 grams; potassiumchloride, 4.0 grams; sodium hydrogen phosphate, 23 grams; potassiumdihydrogen phosphate, 4.0 grams; and optionally phenol red powder, 0.4grams. The solution is sterilized by autoclaving at 15 pounds pressurefor 15 minutes and then diluted with additional water to a singlestrength concentration prior to use.

Alternatively the interferon can be formulated into flavored orunflavored solutions or syrups using a buffered aqueous solution ofinterferon as a base with added caloric or non-caloric sweeteners,flavor oils and pharmaceutically acceptable surfactant/dispersants.

It is also contemplated by the present invention to provide interferonin a solid dosage form such as a lozenge adapted to be dissolved uponcontact with saliva in the mouth with or without the assistance ofchewing. Such a unitary dosage form is formulated to release about 1 toabout 1500 IU of interferon upon dissolution in the mouth for contactwith the oral and pharyngeal mucosa. Thus a unitary dosage form ofinterferon in accordance with this invention can be prepared byart-recognized techniques for forming compressed tablets such aschewable vitamins. Similarly, interferon can be incorporated intostarch-based gel formulations to form a lozenge which will dissolve andrelease interferon for contact with the oral mucosa when held in themouth. Solid unitary dosage forms of interferon for use in accordancewith the present invention can be prepared utilizing art recognizeddosage formulation techniques. The pH of such formulations can rangefrom about 4 to about 8.5. Of course, in processing to such unitarydosage forms one should avoid heating a pre-dosage form formulation,after addition of interferon, above about 50° Centigrade. Exemplary of asolid dosage form for animal use is a molasses block containingeffective amounts of interferon.

Preparation of Human Alpha-Interferon

Human alpha-interferon can be prepared through the following procedure,commonly referred to as the Cantell procedure. The process begins withpacks of human leukocytes, obtained in this case from the Gulf CoastRegional Blood Center, Houston, Texas. The buffy coats in these packsare pooled into centrifuge bottles, and then are diluted with 0.83%ammonium chloride. The mixture is incubated for 15 minutes withintermittent shaking, and is then centrifuged for 20 minutes at 2000rpm. The supernatant is discarded, and the cell pellets are resuspendedwith a minimal volume of sterile phosphate buffered saline (PBS). Themixture is then diluted with ammonium chloride and centrifuged. Thesupernatant is again discarded, and the remaining cell pellets areresuspended with a minimal volume of a tissue culture medium such asMinimal Essential Medium (MEM), available from KC Biological. The cellconcentration is determined with a Coulter counter.

Interferon induction takes place in glass or plastic bottles. Theinduction medium contains MEM, 75 mM Hepes (available from Calbiochem),75 mM Tricine (available from Sigma Chemical Co.), human agamma serum(18 mg/ml), and gentamycin sulfate (from M.A. Bioproducts; 50 mcg/ml).The cells are added to the induction vessels at a final concentration ofabout 5 to 10 million cells per milliliter. The induction vessel isincubated in a 37° C. water bath, and interferon alpha is added as aprimer.

After two hours, Sendai virus is added to the induction mixture. Thiscauses alpha interferon to be produced in the supernatant by theleukocytes. After a 12-18 hour incubation time, the induction mixture iscentrifuged. The cells are discarded, and the supernatant is thenpurified.

The crude interferon is chilled to 10° C. or below in an ice bath. Fivemolar potassium thiocyanate is added to obtain a final concentration of0.5M. This solution is stirred for 15 minutes, and then its pH islowered to 3.3 by adding hydrochloric acid. The mixture is thencentrifuged at 2800 rpm for 30 minutes, and the supernatant isdiscarded.

The pellets are then resuspended in 95% ethanol and are stirred for 15minutes. This suspension is centrifuged at 2800 rpm for 20 minutes, andthe pellets are discarded. The pH of the supernatant is then adjusted to5.8 with sodium hydroxide. The mixture is stirred for 10 minutes, andthen centrifuged at 2800 rpm for 20 minutes. The pellets are discarded.The pH of the supernatant is then adjusted to 8 with sodium hydroxide.This solution is stirred for 10 minutes, followed by centrifugation at2800 rpm for 20 minutes. The supernatant is discarded, and the pelletsare resuspended with 0.5M potassium thiocyanate in a 0.1M sodiumphosphate buffer. This suspension is stirred at 4° C.

Next, the suspension is centrifuged at 2800 rpm for 20 minutes, and thepellets are discarded. The pH of the supernatant is adjusted to 5.3 withhydrochloric acid. After stirring for 10 minutes and centrifugation, thepH of the supernatant is adjusted to 2.8 with hydrochloric acid,followed by further stirring for 20 minutes. This mixture is centrifugedat 2800 rpm, and the resulting pellet is purified humanalpha-interferon.

The pellet is resuspended with 0.5M potassium thiocyanate in 0.1M sodiumphosphate buffer, having a pH of 8.0. It is then dialyzed against PBS at4° C., with two changes of PBS. This mixture is then centrifuged and theprecipitate is discarded. The remaining purified alpha interferon issterilized by filtration through a 0.2 micron filter. A humanalpha-interferon is produced in accordance with this procedure by ImmunoModulators Laboratories, Inc., Stafford, Tex., and sold under thetrademark Agriferon® for use in cattle and Equiferon® for use in horses.

Other procedures known to those skilled in the art are available formaking interferons, such as human alpha-interferon and humangamma-interferon. For example, U.S. Pat. Nos. 4,376,821 and 4,460,685disclose methods of making human gamma-interferon. A method of makingbovine fibroblast (beta) interferon is disclosed in applicant's U.S.Pat. No. 4,462,985.

Clinical Studies

Tables 1-4 below summarize the results of clinical studies of theadministration of interferon by veterinarians orally to 137 dogs andcats as of November, 1985. The studies were conducted with both humanalpha-interferon and bovine beta-interferon. Tables 1-4 compare survivalrates of pets with feline leukemia virus-associated diseases or canineparvovirus disease. Unequal numbers of pets were treated with each typeof interferon; bovine beta-interferon was given to 78 pets and humanalpha-interferon was given to 59 pets.

Bovine beta-interferon was produced in flasks of confluent monolayers ofbovine fetal kidney (BFK) cells. Culture supernatant was harvested 24hours after bluetongue virus induction of BFK cells. The supernatant wasdialyzed 24 hours in a pH 2.0 buffer and for another 24 hours in a PBS(pH.7.4) buffer before interferon assay. Procedures for the assay andcharacterization of bovine beta-interferon were essentially as describedby Rosenquist and Loan, American Journal of Veterinary Research, 28;619-628, 1967. Interferon titers as "units" were expressed as thereciprocals of the dilutions that provided a 50% reduction in the numberof VSV plaques as compared with the number in control cultures. The BFKcell culture interferon produced by this method had an average titer of7,000 units per milliliter. Dogs were given bovine beta-interferon, 5-10ml/dose, as least three times/day after a diagnosis of CPV disease. Catspositive by ELISA for feline leukemia virus and exhibiting clinicalsigns of disease were given 1 ml/10 lb of body weight 2-3 times dailyfor five days. After a five-day interval, cats were retreated at leastonce for another five days.

Human alpha interferon was obtained from IML, Inc. of Houston, Tex.Cases were treated with lot AO26 applied at 6×10⁶ IU/ml. Lot AO26 ofhuman alpha interferon was diluted 1:150 in Eagles' minimum essentialmedium (MEM) and used as the stock solution from which 1 ml was furtherdiluted 1:1000 with 1 liter of MEM for treatment. The usual dose ofhuman alpha interferon was 4 IU/lb body weight given at least threetimes daily after a diagnosis of CPV disease was made. For felineleukemia, cats were treated with human alpha-interferon 2-3 times dailyfor five days as reported for bovine beta-interferon.

Significantly (P<0.05) more cats lived six and twelve months afterdiagnosis and treatment for feline leukemia virus if alpha-interferonwas given, compared to treatment with bovine beta-interferon.Significantly (P<0.05) more dogs survived CPV disease when given alphainterferon (92%) compared to those dogs given bovine beta-interferon(69%).

                  TABLE 1                                                         ______________________________________                                        Summary of Survival Date from clinically ill cats                             positive for FeLV.                                                                       Months After Treatment                                             Treatment    1          6       12                                            ______________________________________                                        Human alpha-IFN                                                                            25/33       21/32*  19/31*                                       Bovine IFN   26/36      15/36   13/36                                         ______________________________________                                         Numerator = no. alive; denominator = no. treated.                             *Cats given human alphaIFN had significantly (P <.05) higher survival         rates at 6 and 12 months after treatment than cats given bovine IFN.          Significance was determined by Chi Square test.                          

                  TABLE 2                                                         ______________________________________                                        Percent survival of clinically ill cats positive for                          FeLV.                                                                                   Months After Treatment                                              Treatment   1            6       12                                           ______________________________________                                        Human alpha-IFN                                                                           76%           66%*    61%*                                        Bovine IFN  72%          42%     36%                                          Historical Control                                                                        <50%         <30%    --                                           ______________________________________                                         Numerator = no. alive; denominator = no. treated.                             *Cats given human alphaIFN had significantly (P <.05) higher survival         rates at 6 and 12 months after treatment than cats given bovine IFN.          Significance was determined by Chi Square test.                          

                  TABLE 3                                                         ______________________________________                                        Response of CPV disease to treatment with bovine                              interferon or human alpha-interferon, by veternarian.                         Attending                                                                              Bovine IFN Beta Human Alpha IFN                                      Veternarian                                                                            Lived     Died      Lived   died                                     ______________________________________                                        S        16/21     5/21      14/16    2/16                                    M        6/11      5/11      7/7     0/7                                      R        7/10      3/10      3/3     0/3                                               29/42 (69%)                                                                             13/42 (31%)                                                                             24/26 (92%)                                                                           2/26 (8%)                                ______________________________________                                    

Dogs treated with human alpha-interferon had a significantly (P<0.05)higher survival rate compared to dogs treated with bovine IFN.Significance between groups was determined by Chi Square test.

                  TABLE 4                                                         ______________________________________                                        Treatment days for CPV disease.                                                          No. of                                                                              Average No. Treatment                                                                        Survival                                      Treatment    Days    Days*     SD**   Rate                                    ______________________________________                                        Bovine IFN   42      3.31      1.95   69%                                     Human alpha IFN                                                                            26      2.75      0.92   92%                                     ______________________________________                                         *Calculated on surviving dogs only.                                           **Standard deviation of the mean treatment days.                         

Canine Herpesvirus Challenge of Newborn Dogs

Canine herpesvirus infection of dogs less than one week of age areinvariably fatal, but older pups usually survive. Interferon has beensuccessfully used to treat viral infections of many species. Thesestudies were conducted to assess the efficacy of interferon in canineherpesvirus (CHV) inoculated puppies.

Five (5) pregnant bitches were obtained from a USDA licensed supplierand were housed in a USDA approved research facility in Canyon, Tex.After the pups whelped, they were inoculated with 6.3 log 10 units ofvirulent CHV obtained from Dr. Richard Mock of the Texas A&M UniversityVeterinary Medical Diagnostic Laboratory (TVMDL) in Amarillo, Tex. Humanalpha-interferon (HAI) or placebo was given to pups orally as treatmentin an effort to increase the survival rate of the CHV inoculated pups.Each litter was divided into control and treated animals. The proceduresand schedule for each litter are discussed below. All dead animals werenecropsied at TVMDL, Amarillo, Tex.

LITTER 1:

Nine (9) pups were inoculated orally with CHV on the day of birth.Interferon was given at 6-10 units (1×), or ten times the dosage at60-100 units (10×). Three pups were given 0.5 ml placebo, 3 pups weregiven 0.5 ml HAI (1×), and 3 pups were given 0.5 ml of a 10× concentrateof HAI orally twice daily for 7 days (if they lived that long). The 3controls died 5, 6, and 8 days after CHV inoculation. The 3 pups givenHAI (1×) lived 7, 7, and 9 days and the 3 pups given 10× HAI lived 6, 7,and 7 days after CHV inoculation.

Pups given HAI (1×) lived an average of 1.3 days longer than controls,but the longer survival time was not statistically significant. Thehigher dosage, HAI (10×), did not provide a survival benefit over thelower dosage, but instead pups given the higher dosage died, on theaverage, one day sooner.

LITTER 2:

Eight (8) pups were inoculated with CHV orally 2 days after birth.Interferon was given at 6-10 units (1×), or ten times the dosage (10×),or 1/10th the dosage (1/10×). All interferon was given orally afterdilution in PBS. Two (2) pups were given 0.5 ml PBS, 2 were given 0.5 mlHAI at 1/10th concentration, 2 were given HAI (1×), and 2 were given a10× concentrate of HAI. All treatments were given orally twice daily for5 days starting 1 day before CHV inoculation. The 2 controls died <1 and9 days after CHV inoculation and the HAI treated (1/10th dose, fulldose, 10× dose) pups died 8 and 9, 5 and 9, 8 and 8 days after CHVinoculation, respectively.

No benefit from treatment at any dosage was seen. The death of a controlpup within a day after CHV inoculation was probably not related to CHVinoculation.

LITTER 3:

Nine (9) pups were inoculated with CHV orally 3 days after birth. Twopups were given 0.5 ml PBS, 2 were given 0.5 ml HAI (1×), 2 were given1/10th dose HAI, and 3 were given 2 IU of recombinant humanalpha-interferon from Schering-Plough. All treatments were given orallytwice daily for 5 days starting two days before CHV inoculation. Bothpups given HAI (1×) survived until necropsied 19 days after CHVinoculation. One control pup died 14 days after CHV inoculation and 1survived until necropsied 19 days after CHV inoculation. Pups given1/10th dosage of HAI died 8 and 13 days after CHV inoculation. Only onepup given recombinant human alpha-interferon died (12 days after CHV);the other 2 pups survived until necropsied 19 days after CHVinoculation.

These pups, inoculated 3 days after birth, did not develop anoverwhelming CHV infection (only 1 of 2 controls died). A 1/10th dose ofHAI did not protect either pup but both HAI (1×) treated pups survived.

LITTER 4:

Fourteen (14) pups were inoculated orally with CHV 2 days after birth.Seven (7) pups were given PBS and 7 pups were given HAI (1×) orallytwice daily for 7 days (if they lived that long) starting 2 days afterCHV inoculation. The 7 controls died 1, 5, 7, 8, 8, 9, and 9 days afterCHV inoculation. One of the HAI (1×) treated pups survived and the other6 pups died 1, 6, 8, 9, 9, and 12 days after CHV inoculation. The deathsof 2 pups only 1 day after CHV inoculation were probably not related toCHV inoculation.

One HAI (1×) treated pup lived 3 days beyond the last surviving controland one HAI (1×) treated pup lived 2 weeks (until necropsied) beyond anytreated control pup. Average survival time of interferon treated pupswas longer than control survival time, but not significantly so.

LITTER 5:

Six (6) pups were inoculated orally with CHV 2 days after birth. Three(3) pups were given PBS and 3 were given HAI (1×) orally once dailystarting 5 days after CHV inoculation. The 3 controls died 6, 6, and 7days after CHV inoculation. One of the HAI (1×) treated pups survived(until necropsied) and the others died 8 and 9 days after CHVinoculation.

All interferon treated pups lived longer than any of the control pups.Treatment with HAI (1×) did not begin until 5 days after CHVinoculation, yet survival was significantly (P<0.05) prolonged.

In summary, on the average, puppies treated with human alpha-interferonhad longer survival times and enhanced survival rates compared tolittermate controls, after canine herpesvirus challenge. A total of 7puppies (1 control and 6 interferon treated) survived the normally fatalCHV inoculation. The data is summarized in the Table 5 below.

                  TABLE 5                                                         ______________________________________                                        Summary of Canine Herpesvirus Data                                                                        Average*                                               No. of                 Survival                                          Litter                                                                             Pups      Dosage       Time (Days)                                                                           Survivors                                 ______________________________________                                        1    3         control      6.33    0                                         1    3         HAI 1× 7.67    0                                         1    3         HAI 10×                                                                              6.67    0                                         2    2         control      4.5**   0                                         2    2         HAI 1/10th   8.5     0                                         2    2         HAI 1× 7.0     0                                         2    2         HAI 10×                                                                              8.0     0                                         3    2         control      14.0    1                                         3    2         HAI 1/10th   10.5    0                                         3    2         HAI 1× --      2                                         3    3         recombinant IFN                                                                            12.0    2                                         4    7         control      6.7     0                                         4    7         HAI 1× 7.5     1                                         5    3         control      6.3     0                                         5    3         HAI 1× 8.5     1                                         ______________________________________                                         *dead dogs survival time; living puppies not calculated.                      **includes one pup living only one day beyond CHV inoculation.           

Treatment Of Nasal Solar Dermatitis

Three cases of nasal solar dermatitis (collie nose) cleared after humanalpha-interferon treatment of 1 unit/lb body weight orally and topicaltreatment (a few ml at 20 units/ml).

Treatment of Canine Lupus Erythematosus

Two cases diagnosed as canine lupus erythematosus were cured by humanalpha-interferon treatment. A 2 year-old Lhasa apso male had beentreated with prednisolone for 1 year for 3 dermatological lesions on theabdomen and prepuce. The flat glistening lesions were continually lickedby the dog. Within 1 week of oral human alpha-interferon treatment (1unit/lb body weight daily for 5 days, then after 1 week, treatment wasrepeated for 5 days) 2 lesions completely healed and the third lesionwas reduced to 1/2 its original size. Within 4 weeks, the lesions wereall completely healed and all therapy ceased. One year later, a skinlesion reappeared but promptly healed after interferon treatment wasrepeated. The skin lesions have not reappeared in the past 10 months.

A 6 year-old spayed female Chihuahua cross had a spider shaped (4 cm by2 cm approximately) skin lesion on the abdomen. The lesion was flat,glistening and pruritic. Six weeks of prednisolone treatment resulted incomplete healing. The following summer, the lesion reappeared and wastreated with oral human alpha-interferon at about 1 unit/lb body weightdaily for 5 days. Within 5 days the lesion was reduced to 1/3 itsoriginal size and completely disappeared within 10 days. The lesion hasnot reappeared in the past year.

Treatment Of Feline Infectious Peritonitis

Table 6 shows the results of 17 cases of feline infections peritonitis(FIP) as diagnosed by practicing veterinarians. Human alpha-interferon(IFN) treatment resulted in a significantly greater survival rate thantreatment with bovine beta-IFN.

                  TABLE 6                                                         ______________________________________                                        Survival of clinically ill cats diagnosed as FIP                                         No. Cats                Survival                                   Treatment  Treated  Alive     Dead Rate                                       ______________________________________                                        Human      11       10        1    91%                                        alpha-IFN                                                                     Bovine      6        3        3    50%                                        beta-IFN                                                                      Total      17       13        4    76%                                        ______________________________________                                    

Cats given human alpha-IFN had a significantly (P=0.0574) greatersurvival rate than cats given bovine beta-IFN.

Human Treatment With Exogenous Human Alpha-Interferon

Human patients were treated with human alpha-interferon in the therapyof acute rheumatoid arthritis, multiple sclerosis, asthma, acne,malignant lymphoma, mesothelioma, and apthous stomatitis. Therapyconsisted of oral administration of 0.7 IU per lb. of patient bodyweight twice daily, once in the morning and once in the evening. None ofthe patients noted any fever or anorexia associated with theadministration of alpha interferon. Interferon was administered in abuffered solution having a concentration such that a single dosage couldbe administered in a volume of about 1 to about 20 ml of liquid. Eachpatient generally retained the interferon solution in his mouth for aperiod of time up to about one minute. After that time the solution waseither swallowed or discharged from the patient's mouth.

Two patients suffering from rheumatoid arthritis were treated--aCaucasian male age 44 and a Caucasian female age 44. The male patientwas pain free in 7 days, and the female was pain free in 10 days. Theywere both continued on the oral interferon for 21 days total and haveremained asymptomatic.

It has been found that recurrence of a treated arthritic condition canbe minimized if treatment in accordance with the present invention iscontinued over a period of up to about three months.

A 30-year-old Caucasian female nurse afflicted with multiple sclerosisand who had had an extensive neurologic workup at City of Hope Hospitalin Los Angeles received treatment in accordance with the presentinvention for 21 days. The patient has had no recurrence of herneurologic symptoms for the past nine months.

A 42-year-old Caucasian male diagnosed to have a malignant lymphoma hadcompleted chemotherapy with dismal results and was considered terminal.He was treated for three weeks with oral interferon. Six months afterstarting treatment he was released by his oncologist as free of thedisease.

An 82-year-old Caucasian female was diagnosed to have mesothelioma.Presently there is no effective treatment for that disease and only a9-month average survival rate is predicted. During her treatment withhuman alpha-interferon she had thoracentesis on two occasions for pluraleffusion. Otherwise, the patient has been active and has survived for 43months.

A 32-year-old Asian male with apthous stomatitis was treated for twoweeks with human alpha-interferon in accordance with the presentinvention. There has been no recurrence of the ulcers over the last sixmonths since treatment was completed.

BKC is a 29 year-old Caucasian female and KKJ is a 20 year-old Caucasianfemale. Both are afflicted by acne-like skin blemishes at the time oftheir monthly menstrual cycle. Oral human alpha-interferon given atabout 1 unit/lb of body weight for 3 days prior to the time of theircycle reduces the severity and number of skin blemishes.

Treatment Of Warts In Humans With Bovine Alpha-Interferon

MAH, a 38 year-old Caucasian Caucasian female, had 7 warts on the middlefinger of her right hand. After 9 months duration, medical treatment wassought, and liquid nitrogen was applied by a dermatologist. Only onewart on the finger regressed after treatment. Three warts coalesced tocreate a large wart area that, over the next year, acquired a roughly 12millimeter square shape. Oral bovine alpha interferon treatment wasstarted at a dosage of 6 ml daily for 6 consecutive days. Theconcentration of alpha-interferon was 30 units/ml; it was derived fromthe nasal secretions of cattle infected with infectious bovinerhinotracheitis virus. All the warts completely regressed within 6 weeksof the first dose of interferon.

Interferon Dosage Formulations

(1) Lozenge

A starch gel-based lozenge containing interferon is prepared bycombining 150 grams of sucrose, 550 ml phosphate buffered saline, and250 grams of a cold-water-soluble starch such as that described in U.S.Pat. No. 4,465,702, heating that mixture with stirring to a temperatureof about 75° C., cooling the mixture to about 30° C. and thereafterblending into the paste-like mass 50 ml of phosphate buffered saline PBScontaining human alpha interferon at a concentration of 250 IU/ml. Themixture is then formed into multiple portions of about 5 to about 10grams each which set upon standing under drying conditions to a starchcandy gel-like consistency. The lozenges thereby produced can beadministered to a patient singly or in combination. The patient isinstructed to hold the lozenge in his mouth until it is completelydissolved to release the interferon component for contact with the oralmucosa.

(2) Chewable Vitamin

A chewable vitamin formulation is prepared, for example, according tothe description of U.S. Pat. No. 3,857,939 by coating one or morecomponents thereof prior to tableting with an interferon solution in anamount sufficient to provide about 1 to about 1500 units of interferonin each chewable vitamin tablet.

(3) Mouthwash

A mouthwash formulation is prepared in accordance with the presentinvention by combining 850 ml PBS, 100 ml of glycerin, 50 grams ofdextrose, and a mixture of 0.3 ml of a flavor oil pre-mixed with 30 mlof a palatable, pharmaceutically acceptable surfactant/dispersant havingan HLB from about 15 to about 25 and 50 ml of a PBS solution ofinterferon (concentration 120 IU/ml). The formulation containsinterferon at a concentration of about 120 IU per 20 ml dosage. Thepatient is asked to hold a 20 ml volume of the mouthwash in his mouth,optionally gargling with the same, for a period of about 15 seconds toabout one minute.

(4) Syrup

Interferon is added to a commercial cough syrup formulation in an amountsufficient to provide an interferon containing syrup formulation havingabout 1 to about 1500 IU of human interferon per tablespoon of syrup.

(5) Effervescent Tablet

A tableting mixture comprising a pharmaceutically acceptable alkalimetal carbonate or bicarbonate, an organic acid such as citric acid,human interferon (preferably dispersed on a suitable organic orinorganic carrier therefor) in an amount sufficient to provide a pertablet dose of about 1 to about 1500 units of interferon per dose, andfurther including suitable tableting excipients such as lubricants andbinders, is compressed into a unitary dosage form of interferon. Thecompressed tablet effervesces upon contact with water to releaseinterferon to the resulting buffered solution. The dosage of interferonis readily available in solution for contact with the oral pharyngealmucosa of a patient in need of said dosage of interferon.

I claim:
 1. A method for treating a human patient afflicted with anautoimmune disorder characterized by chronic inflammatory tissuedegeneration, the method consisting essentially of the stepsof:introducing a saliva-soluble dosage form of interferon into the mouthof the patient, allowing the solid dosage form to dissolve in the mouthto form an interferon-containing solution, and contacting theinterferon-containing solution with the oral and pharyngealmucosa;wherein the interferon is administered to the patient in anamount of about 0.01 to about 5 IU interferon per pound of patient bodyweight per day.
 2. The method of claim 1 wherein the administeredinterferon is human alpha-interferon.
 3. The method of claim 2 whereinthe interferon is administered at a dosage of about 0.1 to about 4.0IU/lb of patient body weight per day.
 4. The method of claim 1 whereinthe autoimmune disorder is selected from the group consisting ofmultiple sclerosis, rheumatoid arthritis, nasal solar dermatitis, andlupus erythematosus.
 5. The method of claim 1 wherein humanalpha-interferon is administered to the human patient at a dosage ofabout 0.5 to about 1.5 IU/lb of patient body weight per day.
 6. Themethod of claim 1 wherein the administered interferon is humanbeta-interferon.
 7. The method of claim 1 wherein the solid dosage formis a lozenge.
 8. The method of claim 2 wherein the solid dosage form isa lozenge.
 9. The method of claim 3 wherein the solid dosage form is alozenge.
 10. The method of claim 4 wherein the solid dosage form is alozenge.
 11. The method of claim 5 wherein the solid dosage form is alozenge.
 12. The method of claim 6 wherein the solid dosage form is alozenge.
 13. A method for treating a human patient afflicted with anautoimmune disorder characterized by chronic inflammatory tissuedegeneration, the method comprising the steps of:introducing asaliva-soluble dosage form of interferon into the mouth of the patient,allowing the solid dosage form to dissolve in the mouth to form aninterferon-containing solution, and contacting the interferon-containingsolution with the oral and pharyngeal mucosa;wherein the interferon isadministered to the patient in an amount of about 0.01 to about 5 IUinterferon per pound of patient body weight per day.
 14. The method ofclaim 13 wherein the interferon is administered at a dosage of about 0.1to about 4.0 IU/lb of patient body weight per day.
 15. The method ofclaim 13 wherein the autoimmune disorder is selected from the groupconsisting of multiple sclerosis, rheumatoid arthritis, nasal solardermatitis, and lupus erythematosus.
 16. The method of claim 13 whereinhuman alpha-interferon is administered to the human patient at a dosageof about 0.5 to about 1.5 IU/lb of patient body weight per day.
 17. Themethod of claim 13 wherein the administered interferon is humanalpha-interferon.
 18. The method of claim 17 wherein the saliva solubledosage form is a lozenge.